# 1360: HIPPEA JASONIAE MEDIUM


## Main sol. 1360

26.33	g	NaCl	
1.5	g	MgCl2 x 6 H2O	
0.45	g	KCl	
1.3	g	(NH4)2SO4	
0.28	g	KH2PO4	
0.25	g	MgSO4 x 7 H2O	
0.07	g	CaCl2 x 2 H2O	
0.02	g	FeCl3 x 6 H2O	
10	ml	**Allen's trace element solution**	
1.25	g	Na-acetate	
2.94	g	Trisodium citrate x 2 H2O	
0.5	ml	Sodium resazurin (0.1% w/v)	
10	g	Sulfur, powdered	
0.5	g	Yeast extract (DIFCO)	
1	ml	**Wolin's vitamin solution (10x)**	
0.5	g	Na2S x 9 H2O	
1000	ml	Distilled water	

1. Dissolve ingredients (except sulfur, vitamins, yeast extract and sulfide), adjust pH to 3.5 with 10 N H2SO4 and sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into serum vials (e.g., 20 ml medium in 100 ml bottles) that contain already the appropriate amount of sulfur. Sterilize medium by heating cultivation vessels in a water bath to 90 - 100°C for 1 - 2 hours on each of 3 successive days. Add vitamins (sterilized by filtration) and yeast extract from sterile anoxic stock solutions prepared under 100% N2 gas. Prior to inoculation, add sulfide from a sterile anoxic stock solution prepared under 100% N2 gas and adjust pH of complete medium to 4.5, if necessary.


## Allen's trace element solution

180	mg	MnCl2 x 4 H2O	
450	mg	Na2B4O7 x 10 H2O	
22	mg	ZnSO4 x 7 H2O	
5	mg	CuCl2 x 2 H2O	
3	mg	Na2MoO4 x 2 H2O	
3	mg	VOSO4 x 2 H2O	
1	mg	CoSO4 x 7 H2O	
1000	ml	Distilled water	

1. Adjust pH of final solution to 2 with 1 N HCl.


## Wolin's vitamin solution (10x)

20	mg	Biotin	
20	mg	Folic acid	
100	mg	Pyridoxine hydrochloride	
50	mg	Thiamine HCl	
50	mg	Riboflavin	
50	mg	Nicotinic acid	
50	mg	Calcium D-(+)-pantothenate	
1	mg	Vitamin B12	
50	mg	p-Aminobenzoic acid	
50	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water